Ovarian cancers are highly lethal tumors which account for approximately four percent of all women's cancers and are the fifth leading cause of cancer-related death among women.
Ovarian cancer is defined as cancer that forms in the tissues of the ovary (National Cancer Institute. (n.d.) Ovarian Cancer). Most ovarian cancers start in either the cells on the surface of the ovary (epithelial carcinoma) or in the egg cells themselves (germ cell tumors) (National Cancer Institute. (n.d.) Ovarian Cancer). According to the Centers for Disease Control and Prevention, ovarian cancer is responsible for more deaths among women than any other cancer of the reproductive system (Centers for Disease Control and Prevention. (2014, March) Ovarian Cancer). There have been 14,270 cases of ovarian cancer in 2014 alone (National Cancer Institute. (n.d.) Ovarian Cancer). One of every 68 women will develop ovarian cancer in their lifetime (Tung, C H et al (2014) Lessons learned from imaging mouse ovarian tumors: the route of probe injection makes a difference. Quant Imaging Med Surg, 4(3):156-162). Early diagnosis of ovarian cancer increases the chances of survival but only 14.7% of ovarian cancers are diagnosed in the local stage where the cancer has not spread outside the ovary (National Cancer Institute. (n.d.) Cancer Statistics).
The difficulty in catching ovarian cancers early lies in the fact that there may be no symptoms or the symptoms may be very common such as bloating and abdominal pain. Women with a family history of ovarian cancer or between the ages of 55 and 64 are most frequently diagnosed with ovarian cancer (National Cancer Institute. (n.d.) Cancer Statistics). Treatments for ovarian cancer include cytoreductive surgery and chemotherapy (Khabele, D (2014) The Therapeutic Potential of Class I Selective Histone Deacetylase Inhibitors in Ovarian Cancer. Front Oncol, 4; 4: 111). Cytoreductive surgery aims to debulk the tumor while platinum-based chemotherapy serves as a systemic therapy. Despite the treatment options available, women with advanced stages of ovarian cancer have low chances of survival.
Ovarian cancer patients often initially respond well to the platinum-based chemotherapy but eventually experience lower survival outcomes due to chemotherapy-induced resistance which can often occur rapidly and become fatal (Echevarría-Vargas I M, et al (2014) Upregulation of miR-21 in Cisplatin Resistant Ovarian Cancer via JNK-1/c-Jun Pathway. PLoS ONE 9 (5)).
At diagnosis the majority of patients have metastatic disease and the long-term survival remains low. Certain ovarian cancers are highly lethal tumors due to the emergence of therapy-resistant ovarian cancer cells.
Protein Kinase C (PKC) and Ovarian Cancer
The protein kinase C (PKC) family of Ser/Thr kinases is involved in transmembrane signal transduction pathways triggered by various extra and intracellular stimuli (Nelson D L Cox M M. Biosignaling. Lehninger Principles of Biochemistry. 3rd ed. New York: Worth Publishers; 2001. p. 469). They are involved in the control of cellular responses that include proliferation, migration, apoptosis and survival (Couldwell W T, Antel J P, Apuzzo M L, Yong V W. Inhibition of growth of established human glioma cell lines by modulators of the protein kinase-C system. J Neurosurg 1990; 73:594-0; Okhrimenko H, Lu W, Xiang C, Hamburger N, Kazimirsky G, Brodie C. Protein Kinase C-ε Regulates the Apoptosis and Survival of Glioma Cells. Cancer Res 2005; 65:7301-7309; Stensman H, Larsson C. Protein kinase C epsilon is important for migration of neuroblastoma cells. BMC Cancer 2008; 8:365). PKC regulates cellular functions, metabolism and proliferation by phosphorylating proteins in response to transmembrane signals from hormones, growth factors, neuro-transmitters and pharmacological agents.
Protein kinase C (PKC) is a family of fourteen known isozymes found in varying ratios in the cytosolic and membrane fractions of cells, depending on the type of tissue and its physiological state (Nishizuka 1992 Science 258, 607). PKC isozymes can be classified into three groups. Group I includes Ca2+ dependent isozymes: cPKC-alpha, cPKC-betaI cPKC-betaII and cPKC-gamma. Isozymes in group II, nPKC-epsilon, nPKC-delta, nPKC-eta and nPKC-theta are Ca2+ independent. Group III includes the atypical PKC: aPKC-iota (Selbie et al. 1993 J. Biol. Chem. 268, 24296), aPKC-zeta, aPKC-zetaII (Hirai et al. 2003 Neuroscience Lett. 348, 151), aPKC-mu (protein kinase D) and aPKC-nu (Hayashi et al. 1999 Biochim. et Biophys. Acta. 1450, 99) which are insensitive to both diacylglycerol and calcium and neither bind to nor are activated by phorbol esters. PKC-ζ and PKC-ι exhibit 72% sequence homology at the amino acid level. This structural similarity coupled with the fact that many commercial immunological reagents do not distinguish between these isoforms, has made it difficult to biochemically distinguish between PKC-ζ and PKC-ι. (Fields A P, Regala R P. Protein kinase Cι: Human oncogene, prognostic marker and therapeutic target. Pharmacol Res 2007; 55:487-97)
Protein kinase C-iota (PKC-ι) has been shown to aid in the ability of cancer cells to resist drug-induced apoptosis. Recently, it has been reported that PKC-ι which is located in chromosome 3 at 3q26.2 is the most common genomic amplicon as identified by comparative genomic hybridization (Eder A M, Sui X, Rosen D G, Nolden L K, Cheng K W, Lahad J P, Kango-Singh M, Lu K H, Warneke C L, Atkinson E N, Bedrosian I, Keyomarsi K, Kuo W L, Gray J W, Yin J C, Liu J, Halder G, Mills G B. Atypical PKC iota contributes to poor prognosis through loss of apical-basal polarity and cyclin E overexpression in ovarian cancer. Proc Natl Acad Sci USA 102:12519-12524 (2005)).
PKC-ι protein is markedly increased or mislocalized in all serous ovarian cancers. In nonserous ovarian cancers, increased PKC-ι protein levels, particularly in the presence of Cyclin E, are associated with markedly decreased overall survival. Additionally, an increase in PKC-ι DNA copy number was associated with decreased progression-free survival of ovarian cancer patients. (Eder, A. et al., Atypical PKC-ι contributes to poor prognosis through loss of apical-basal polarity and Cyclin E overexpression in ovarian cancer, PNAS, 2005, 102(35):12519-12524). Moreover, only PKC-ι gene amplification is highly correlated with protein overexpression, tumor size, lymph node metastasis and clinical stage out of four genes studied on the 3q26 amplification [Genes Chromosomes Cancer 47:127-136 (2008)].
Like other PKC isoforms, protein kinase-C zeta (PKC-ζ) is a serine/threonine kinase that adds phosphate groups to target proteins. It is atypical in that unlike other PKC isoforms, PKC-ζ does not require calcium or diacylglycerol (DAG) to become active, but rather relies on a different second messenger, presumably generated through a phosphoinositide 3-kinase (PI3-kinase) pathway. Protein kinase-C zeta (PKC-ζ) has been widely implicated in the regulation of cellular functions such as being a key regulator of critical intracellular signaling pathways induced by various extracellular stimuli. Studies have demonstrated the involvement of PKC-ζ in the mitogen-activated protein kinase cascade, transcriptional factor NFkB activation, ribosomal S6-protein kinase signaling, and cell polarity. An important molecular event in a cell is the association of PKC-ζ with other signaling molecules, as well as scaffold proteins, to form large complexes that regulate their pathways. (Hirai T. et al., Protein Kinase Czeta (PKCzeta): activation mechanisms and cellular functions, J. Biochem., (2003), 133(1):1-7).
PKC-ζ has been shown to be upregulated in ovarian carcinomas. It was found that expression of PKC-ζ in normal surface ovarian epithelial cells and in cystadenomas is absent or very low. However, most human ovarian adenocarcinomas expressed high amounts of PKC-ζ, which correlated with poor prognosis. (Nazarenko I., et al., Atypical Protein Kinase C Zeta Exhibits a Proapoptotic Function in Ovarian Cancer, Molecular Cancer Research, 2010, 8 (6):919-34) Recent studies have supported that PKC-ζ is a potential regulatory component of the IGF1R and ITGB3 pathways and may have a critical role in ovarian tumorgenesis. Researchers found that up-regulation of PKC-ζ leads to expression alterations of IGF1R and ITGB3 in SKOV3 and OVCAR3 cell lines, suggesting that PKC-ζ may participate in ovarian cancer progression by modulating the expression of other important signaling molecules. Further, an increase in cell proliferation in SKOV3 cells was shown when PKC-ζ was over-expressed and SKOV3 cells exhibited a decrease in cell migration when endogenous PKC-ζ expression was down-regulated by small-interference RNA (siRNA). (Seto, K., et al., Atypical Protein Kinase C Zeta: Potential Player in Cell Survival and Cell Migration of Ovarian Cancer, PLoS One, 2015, 10(4):e)123528).
PKC Inhibitors
PKC inhibitors range in their selectivity for a particular class of PKCs. It has been suggested that only atypical isoforms of the PKC family contain the PB 1 domain and thus agents that disrupt signaling through this mechanism should be specific for atypical PKCs. Sodium aurothiomalate (ATM) (1) and aurothioglucose (ATG) (2) both seem to bind in the low micromolar range to PKC-ι, and as such, may be alternatives for PKC-ι specific inhibitors. Another gold-containing drug, auranofin (3), has a similar structure and thus may also have potential as a PKC inhibitor. Structures for the gold-containing inhibitors are shown below. (Roffey, J. and Ott, G. Section editors, Modulators of Atypical Protein Kinase C as Anticancer Agents in Annual Reports in Medicinal Chemistry, Desai, M. editor, Academic Press, 2014, Vol. 49:189-202)

PKC-iota inhibitor [4-(5-amino-4-carbamoylimidazol-1-yl)-2,3-dihydroxycyclopentyl]methyl dihydrogen phosphate, known as (ICA-1) (4), is a small molecule inhibitor which binds to the catalytic domain of human PKC-ι, at amino acid residues 469-475 (glutamine-469, isoleucine-470, arginine-471, isoleucine-472, proline-473, arginine-474, serine-475). ICA-1 is an inhibitory agent specific to PKC-ι. Structure of PKC-ι is shown below.

2-acetyl-1,3-cyclopentanedione (ACPD) is a pan-aPKC inhibitor which inhibits both PKC-ζ and PKC-ι, but not PKC-α, PKC-β, PKC-δ, or PKC-ε. The structure of ACPD is shown below:

Pachastrissamine and its stereoisomers (6-10) have demonstrated cytotoxic effects on several cancer cell lines with potencies in the submicromolar range. Complete inhibition of both PKC-ζ and PKC-ι at 10 μM has been observed with only modest effects for novel and classical PKC isoforms. Screening at lower concentrations showed about 50% inhibition at 3 μM and no inhibition at 1 μM. Structures for pachastrissamine and its stereoisomers are shown below. (Roffey 2014)

A 13-mer PKC-ζ inhibitory peptide (ZIP) (11) is myristoylated at the N-terminus to improve cell penetration and may serve as a novel PKC-ζ specific therapeutic. Structure of ZIP is shown below. (Roffey 2014)

A series of speciosterosulfates (sterolsulfates) (20-23), isolated from the marine sponge Spheciospongia, have been found to inhibit PKC-ζ. Structures for speciosterosulfates are shown below. (Roffey 2014)

Several allosteric inhibitors (12-14) which bind the PIF-1 site and regulate activity through the C1 domain as well as a series of phenylthiopenes (15-17) may also have implications as potential aPKC inhibitors. Structures for the allosteric inhibitors (12-14) and the phenylthiopenes (15-17) are shown below. (Roffey 2014)

Other potential PKC inhibitors include, but are not limited to: a compound containing a maleimide substructure (19); a series of compounds having an indazole-benzimidazole motif (24-25); a series of ATP-competitive thieno[2,3-d]pyrimidine analogues (26-27); a pyrrole amide PKCzI257.3 (28); and a series of 3-hydroxy-2-(3-hydroxyphenyl)-4H-1-benzopyran-4-ones which may bind to the ATP-cleft of the kinase through a keto-hydroxyl motif in multiple conformations (30-33). Structures for the above listed potential PKC inhibitors are shown below. (Roffey 2014)

In light of the current difficulties in ovarian cancer treatment, particularly the emergence of therapy-resistant ovarian cancer cells, what is needed is a new method of treating ovarian cancer which overcomes the obstacles of the prior art.